scholarly journals Involvement of MexS and MexEF-OprN in Resistance to Toxic Ion Chelators in Pseudomonas putida KT2440

2020 ◽  
Vol 8 (11) ◽  
pp. 1782
Author(s):  
Tania Henriquez ◽  
Tom Baldow ◽  
Yat Kei Lo ◽  
Dina Weydert ◽  
Andreas Brachmann ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Efflux Pump ◽  
Toxic Metal ◽  
Growth Defect ◽  
Metal Chelators ◽  
Pseudomonas Putida Kt2440 ◽  
Pseudomonas Species ◽  
Resistance Nodulation Division ◽  
Rnd Efflux Pump

Bacteria must be able to cope with harsh environments to survive. In Gram-negative bacteria like Pseudomonas species, resistance-nodulation-division (RND) transporters contribute to this task by pumping toxic compounds out of cells. Previously, we found that the RND system TtgABC of Pseudomonas putida KT2440 confers resistance to toxic metal chelators of the bipyridyl group. Here, we report that the incubation of a ttgB mutant in medium containing 2,2’-bipyridyl generated revertant strains able to grow in the presence of this compound. This trait was related to alterations in the pp_2827 locus (homolog of mexS in Pseudomonas aeruginosa). The deletion and complementation of pp_2827 confirmed the importance of the locus for the revertant phenotype. Furthermore, alteration in the pp_2827 locus stimulated expression of the mexEF-oprN operon encoding an RND efflux pump. Deletion and complementation of mexF confirmed that the latter system can compensate the growth defect of the ttgB mutant in the presence of 2,2’-bipyridyl. To our knowledge, this is the first report on a role of pp_2827 (mexS) in the regulation of mexEF-oprN in P. putida KT2440. The results expand the information about the significance of MexEF-OprN in the stress response of P. putida KT2440 and the mechanisms for coping with bipyridyl toxicity.

Tigecycline efflux in Acinetobacter baumannii is mediated by TetA in synergy with RND-type efflux transporters

2020 ◽  
Vol 75 (5) ◽  
pp. 1135-1139 ◽  
Author(s):  
Wuen Ee Foong ◽  
Jochen Wilhelm ◽  
Heng-Keat Tam ◽  
Klaas M Pos
Keyword(s):  
Acinetobacter Baumannii ◽  
Efflux Pump ◽  
Gene Knockout ◽  
Drug Susceptibility ◽  
Major Facilitator Superfamily ◽  
Rt Pcr ◽  
E Coli ◽  
Rnd Efflux Pump ◽  
Major Facilitator

Abstract Objectives To investigate the role of Major Facilitator Superfamily (MFS)-type transporters from Acinetobacter baumannii AYE in tigecycline efflux. Methods Two putative tetracycline transporter genes of A. baumannii AYE (tetA and tetG) were heterologously expressed in Escherichia coli and drug susceptibility assays were conducted with tigecycline and three other tetracycline derivatives. The importance of TetA in tigecycline transport in A. baumannii was determined by complementation of tetA in WT and Resistance Nodulation cell Division (RND) gene knockout strains of A. baumannii ATCC 19606. Gene expression of the MFS-type tetA gene and RND efflux pump genes adeB, adeG and adeJ in A. baumannii AYE in the presence of tigecycline was analysed by quantitative real-time RT–PCR. Results Overproduction of TetA or TetG conferred resistance to doxycycline, minocycline and tetracycline in E. coli. Cells expressing tetA, but not those expressing tetG, conferred resistance to tigecycline, implying that TetA is a determinant for tigecycline transport. A. baumannii WT and RND-knockout strains complemented with plasmid-encoded tetA are significantly less susceptible to tigecycline compared with non-complemented strains. Efflux pump genes tetA and adeG are up-regulated in A. baumannii AYE in the presence of subinhibitory tigecycline concentrations. Conclusions TetA plays an important role in tigecycline efflux of A. baumannii by removing the drug from cytoplasm to periplasm and, subsequently, the RND-type transporters AdeABC and AdeIJK extrude tigecycline across the outer membrane. When challenged with tigecycline, tetA is up-regulated in A. baumannii AYE. Synergy between TetA and the RND-type transporters AdeABC and/or AdeIJK appears necessary for A. baumannii to confer higher tigecycline resistance via drug efflux.

scholarly journals Mechanisms of Resistance to Chloramphenicol in Pseudomonas putida KT2440

2011 ◽  
Vol 56 (2) ◽  
pp. 1001-1009 ◽  
Author(s):  
Matilde Fernández ◽  
Susana Conde ◽  
Jesús de la Torre ◽  
Carlos Molina-Santiago ◽  
Juan-Luis Ramos ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Efflux Pump ◽  
Protein Biosynthesis ◽  
Cellular Stress Response ◽  
Knockout Mutant ◽  
Pseudomonas Putida Kt2440 ◽  
Altered Expression ◽  
Content Type ◽  
Reading Frame ◽  
A Genome

ABSTRACTPseudomonas putidaKT2440 is a chloramphenicol-resistant bacterium that is able to grow in the presence of this antibiotic at a concentration of up to 25 μg/ml. Transcriptomic analyses revealed that the expression profile of 102 genes changed in response to this concentration of chloramphenicol in the culture medium. The genes that showed altered expression include those involved in general metabolism, cellular stress response, gene regulation, efflux pump transporters, and protein biosynthesis. Analysis of a genome-wide collection of mutants showed that survival of a knockout mutant in the TtgABC resistance-nodulation-division (RND) efflux pump and mutants in the biosynthesis of pyrroloquinoline (PQQ) were compromised in the presence of chloramphenicol. The analysis also revealed that an ABC extrusion system (PP2669/PP2668/PP2667) and the AgmR regulator (PP2665) were needed for full resistance toward chloramphenicol. Transcriptional arrays revealed that AgmR controls the expression of thepqqgenes and the operon encoding the ABC extrusion pump from the promoter upstream of open reading frame (ORF) PP2669.

scholarly journals Emergence and molecular basis of azithromycin resistance in typhoidal Salmonella in Dhaka, Bangladesh

2019 ◽  
Author(s):  
Yogesh Hooda ◽  
Senjuti Saha ◽  
Mohammad S I Sajib ◽  
Hafizur Rahman ◽  
Stephen P Luby ◽  
...  
Keyword(s):  
Efflux Pump ◽  
Single Point ◽  
Salmonella Typhi ◽  
Oral Drug ◽  
Single Point Mutation ◽  
E Coli ◽  
Vaccine Introduction ◽  
Rnd Efflux Pump ◽  
Azithromycin Resistance

With rising fluoroquinolone and ceftriaxone-resistant Salmonella Typhi, azithromycin, a macrolide, has become the last oral drug available against typhoid. Between 2009-2016, we isolated 1,082 Salmonella Typhi and Paratyphi A strains in Bangladesh, 13 (12 Typhi and 1 Paratyphi A) of which were azithromycin-resistant. When compared to 462 previously sequenced Typhi strains, the genomes of the 12 azithromycin-resistant Typhi strains (4.3.1 sub-clade, H58) harbored an exclusive non-synonymous single-point mutation R717Q in AcrB, an RND-efflux pump. Expression of AcrB-R717Q in E. coli and Typhi strains increased its minimum inhibitory concentration (MIC) for azithromycin by 11- and 3-fold respectively. The azithromycin-resistant Paratyphi A strain also contained a mutation at R717 (R717L), whose introduction in E. coli and Paratyphi A strains increased MIC by 7- and 3-fold respectively, confirming the role of R717 mutations in conferring azithromycin resistance. With increasing azithromycin use, strains with R717 mutations may spread leading to treatment failures, making antibiotic stewardship and vaccine introduction imperative.

scholarly journals Metabolic role of aldehyde dehydrogenases in Pseudomonas putida KT2440

2020 ◽  
Vol 34 (S1) ◽  
pp. 1-1
Author(s):  
Adriana Julián-Sánchez ◽  
Adeli P. Castrejón-Gonzaga ◽  
Gabriel Moreno-Hagelsieb ◽  
Rosario A. Muñoz-Clares ◽  
Héctor Riveros-Rosas
Keyword(s):  
Pseudomonas Putida ◽  
Pseudomonas Putida Kt2440 ◽  
Aldehyde Dehydrogenases ◽  
Metabolic Role

scholarly journals Raising the stakes: Loss of efflux-pump regulation decreases meropenem susceptibility in Burkholderia pseudomallei

2017 ◽  
Author(s):  
Derek S. Sarovich ◽  
Jessica R. Webb ◽  
Matthew C. Pitman ◽  
Linda T. Viberg ◽  
Mark Mayo ◽  
...  
Keyword(s):  
Burkholderia Pseudomallei ◽  
Efflux Pump ◽  
Fatal Outcome ◽  
Antibiotic Administration ◽  
Over Expression ◽  
Resistance Nodulation Division ◽  
Life Threatening ◽  
Rnd Efflux Pumps ◽  
Rnd Efflux Pump ◽  
Repeated Infections

AbstractBurkholderia pseudomallei, the causative agent of the high-mortality disease melioidosis, is a Gram-negative bacterium that is naturally resistant to many antibiotics. There is no vaccine for melioidosis, and effective eradication is reliant on biphasic and prolonged antibiotic administration. The carbapenem drug, meropenem, is the current gold-standard option for treating severe melioidosis. Intrinsic B. pseudomallei resistance towards meropenem has not yet been documented; however, resistance could conceivably develop over the course of infection, leading to prolonged sepsis and treatment failure. Here, we document 11 melioidosis cases in which B. pseudomallei isolates developed decreased susceptibility towards meropenem during treatment, including two cases not treated with this antibiotic. Meropenem minimum inhibitory concentrations increased over time from 0.5-0.75 to 3-8 μg/mL. Using comparative genomics, we identified multiple mutations affecting multidrug resistance-nodulation-division (RND) efflux pump regulators, leading to over-expression of their corresponding pumps. The most commonly affected pump was AmrAB-OprA, although alterations in the local regulators of BpeEF-OprC or BpeAB-OprB were observed in three cases. This study confirms the role of RND efflux pumps in decreased meropenem susceptibility in B. pseudomallei. Further, we document two concerning examples of severe melioidosis where the reduced treatment efficacy of meropenem was associated with a fatal outcome.Significance StatementThe bacterium Burkholderia pseudomallei, which causes the often-fatal tropical disease melioidosis, is difficult to eradicate. Due to high levels of intrinsic antibiotic resistance, only a handful of antibiotics are effective against this pathogen. One of these, meropenem, is commonly used in the treatment of melioidosis patients who are unresponsive to other treatments or are critically ill. Here, we describe 11 melioidosis cases whereby patients exhibited prolonged or repeated infections that were associated with the development of decreased meropenem susceptibility. We identified the molecular basis for this decreased susceptibility in latter B. pseudomallei isolates obtained from these patients, and functionally confirmed the mechanism conferring this phenotype. Our findings have important ramifications for the diagnosis, treatment and management of life-threatening melioidosis cases.

The role of RND efflux pump and global regulators in tigecycline resistance in clinicalAcinetobacter baumanniiisolates

2015 ◽  
Vol 10 (3) ◽  
pp. 337-346 ◽  
Author(s):  
Henan Li ◽  
Xiaojuan Wang ◽  
Yawei Zhang ◽  
Chunjiang Zhao ◽  
Hongbin Chen ◽  
...  
Keyword(s):  
Efflux Pump ◽  
Global Regulators ◽  
Rnd Efflux Pump

Addressing the role of the extrusion pump-bearing pGRT1 plasmid in toluene biodegradation by Pseudomonas putida DOT-T1E under real case scenarios

2009 ◽  
Vol 60 (9) ◽  
pp. 2391-2398 ◽  
Author(s):  
M. Hernandez ◽  
J. Gouveia ◽  
A. Segura ◽  
R. Muñoz ◽  
S. Villaverde
Keyword(s):  
Pseudomonas Putida ◽  
Wild Type Strain ◽  
Efflux Pump ◽  
Similar Process ◽  
Process Performance ◽  
Wild Type ◽  
Mediated Effects ◽  
Genes Encoding ◽  
Toluene Biodegradation

The role of both the plasmid pGRT1 and the solvent extrusion pump ttgGHI during toluene biodegradation in Pseudomonas putida DOT-T1E cultures was investigated in a sterile suspended growth bioreactor operated as chemostat and inoculated independently with a wild type strain, a mutant lacking the pGRT1 plasmid (P. putida DOT-TIE-100), and a mutant with a modified pGRT1 plasmid lacking the genes encoding the ttgGHI solvent efflux pump (P. putida DOT-TIE-28). A similar process performance was recorded in all tested strains at 4 g tol m−3 and dilution rates (D) of 0.1 h−1. However, operation at 10 g tol m−3 and D of 0.2 h−1 revealed a much lower toluene EC (285 g m−3 h−1) in P. putida DOT-T1E-100 cultures when compared to wild type and P. putida DOT-T1E-28 cultures (483–498 g m−3 h−1), which suggests that other mechanisms rather than solvent extrusion by the ttgGHI efflux pump supported this superior process performance. Finally, the plasmid pGRT1 analysed exhibited a remarkable stability towards toluene harmful mediated effects, regardless the strain or toluene loading tested.

scholarly journals Novel Toxin-Antitoxin Module SlvT-SlvA Regulates Megaplasmid Stability and Incites Solvent Tolerance in Pseudomonas putida S12

2020 ◽  
Vol 86 (13) ◽  
Author(s):  
Hadiastri Kusumawardhani ◽  
David van Dijk ◽  
Rohola Hosseini ◽  
Johannes H. de Winde
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Putida ◽  
Aromatic Compounds ◽  
Efflux Pump ◽  
Gene Clusters ◽  
Host Cells ◽  
Solvent Tolerance ◽  
Content Type ◽  
Solvent Tolerant

ABSTRACT Pseudomonas putida S12 is highly tolerant of organic solvents in saturating concentrations, rendering this microorganism suitable for the industrial production of various aromatic compounds. Previous studies revealed that P. putida S12 contains the single-copy 583-kbp megaplasmid pTTS12. pTTS12 carries several important operons and gene clusters facilitating P. putida S12 survival and growth in the presence of toxic compounds or other environmental stresses. We wished to revisit and further scrutinize the role of pTTS12 in conferring solvent tolerance. To this end, we cured the megaplasmid from P. putida S12 and conclusively confirmed that the SrpABC efflux pump is the major determinant of solvent tolerance on the megaplasmid pTTS12. In addition, we identified a novel toxin-antitoxin module (proposed gene names slvT and slvA, respectively) encoded on pTTS12 which contributes to the solvent tolerance phenotype and is important for conferring stability to the megaplasmid. Chromosomal introduction of the srp operon in combination with the slvAT gene pair created a solvent tolerance phenotype in non-solvent-tolerant strains, such as P. putida KT2440, Escherichia coli TG1, and E. coli BL21(DE3). IMPORTANCE Sustainable alternatives for high-value chemicals can be achieved by using renewable feedstocks in bacterial biocatalysis. However, during the bioproduction of such chemicals and biopolymers, aromatic compounds that function as products, substrates, or intermediates in the production process may exert toxicity to microbial host cells and limit the production yield. Therefore, solvent tolerance is a highly preferable trait for microbial hosts in the biobased production of aromatic chemicals and biopolymers. In this study, we revisit the essential role of megaplasmid pTTS12 from solvent-tolerant Pseudomonas putida S12 for molecular adaptation to an organic solvent. In addition to the solvent extrusion pump (SrpABC), we identified a novel toxin-antitoxin module (SlvAT) which contributes to short-term tolerance in moderate solvent concentrations, as well as to the stability of pTTS12. These two gene clusters were successfully expressed in non-solvent-tolerant strains of P. putida and Escherichia coli strains to confer and enhance solvent tolerance.

scholarly journals Pumping iron to keep fit: modulation of siderophore secretion helps efficient aromatic utilization in Pseudomonas putida KT2440

Microbiology ◽  
2014 ◽  
Vol 160 (7) ◽  
pp. 1393-1400 ◽  
Author(s):  
Hiren Joshi ◽  
Rachna Dave ◽  
V. P. Venugopalan
Keyword(s):  
Pseudomonas Putida ◽  
Compensatory Mechanism ◽  
Limited Information ◽  
Pseudomonas Putida Kt2440 ◽  
Physiological Factors ◽  
Aromatic Substrates ◽  
Iron Assimilation ◽  
Key Factor ◽  
Alternative Carbon Source

Studies of biotechnology applications of Pseudomonas putida KT2440 have been predominantly focused on regulation and expression of the toluene degradation (TOL) pathway. Unfortunately, there is limited information on the role of other physiological factors influencing aromatic utilization. In this report, we demonstrate that P. putida KT2440 increases its siderophore secretion in response to the availability of benzyl alcohol, a model aromatic substrate. It is argued that accelerated siderophore secretion in response to aromatic substrates provides an iron ‘boost’ which is required for the effective functioning of the iron-dependent oxygenases responsible for ring opening. Direct evidence for the cardinal role of siderophores in aromatic utilization is provided by evaluation of per capita siderophore secretion and comparative growth assessments of wild-type and siderophore-negative mutant strains grown on an alternative carbon source. Accelerated siderophore secretion can be viewed as a compensatory mechanism in P. putida in the context of its inability to secrete more than one type of siderophore (pyoverdine) or to utilize heterologous siderophores. Stimulated siderophore secretion might be a key factor in successful integration and proliferation of this organism as a bio-augmentation agent for aromatic degradation. It not only facilitates efficient aromatic utilization, but also provides better opportunities for iron assimilation amongst diverse microbial communities, thereby ensuring better survival and proliferation.

Sign in / Sign up

Export Citation Format

Share Document